Sequence and characterization of phocine interleukin 2.
To improve assessment of cellular immune responses in seals, northern elephant seal (Mirounga angustirostris) interleukin 2 (IL-2) has been characterized. The gene was cloned and sequenced from a 658 base pair (bp) cDNA generated from total RNA by reverse transcription-polymerase chain reaction (RT-PCR). The sequence encoded a 154 amino acid (aa) polypeptide that included a 20 aa putative signal peptide. Seal IL-2 was found to share considerable identity with published sequences. Nucleotide sequence analysis of phocine (seal) IL-2 with canine, feline, human, trichechine (manatee), bovine and murine sequences demonstrated 93, 92, 86, 82, 78 and 71% identity, respectively. Analysis of the derived amino acid sequences demonstrated 88, 89, 78, 71, 66 and 60% identity, respectively. Interleukin-2 sequence identities appear to reflect evolutionary proximity among the analyzed species, and importantly, those residues identified as critical to IL-2 biological activity and receptor binding are largely conserved. To examine the kinetics of IL-2 mRNA expression, northern elephant seal lymphocytes were stimulated with the mitogen concanavalin A (Con A), and RNA was collected at several time points thereafter. The RT-PCR demonstrated that seal IL-2 mRNA expression peaks in the first 8 hr following Con A stimulation. Lastly, genomic DNA from northern elephant seal, harbour seal (Phoca vitulina) and California sea lion (Zalophus californianus) was used as template to identify and clone genomic IL-2. Partial sequence of the genomic clones demonstrated nearly complete identity among the three species. Sequence identity indicates that probes constructed from the northern elephant seal IL-2 gene will be effective in assessing IL-2 in other pinniped species.
Shoda
L K
LK
Department of Wildlife and Fisheries Sciences, Texas A&M University, College Station 77843-2258, USA.
Brown
W C
WC
Rice-Ficht
A C
AC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017124:Conserved Sequence; D003429:Cross Reactions; D004247:DNA / Q000737:chemistry; D015870:Gene Expression; D007376:Interleukin-2 / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D007700:Kinetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D046048:Seals, Earless / Q000235:genetics / Q000276:immunology*; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid
D007376:Interleukin-2; D012333:RNA, Messenger; D004247:DNA